Joshua Kogot

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Bacterial Display Peptides for Use in Biosensing Applications

Peptide affinity reagent technology including bacterial display encapsulates an in vitro analog o... more Peptide affinity reagent technology including bacterial display encapsulates an in vitro analog of classical Darwinian evolution through a physical linkage between polypeptide sequence and the encoding of genetic information. Similar to the inate human imune system a unique binder is isolated, however, in this directed evolution method a large library

Screening of Peptide Libraries against Protective Antigen of Bacillus anthracis in a Disposable Microfluidic Cartridge

Rapid Peptide Reagent Isolation in a Disposable Microfluidic Cartridge

: In this collaborative development project, the challenges in affinity reagent isolation were ad... more : In this collaborative development project, the challenges in affinity reagent isolation were addressed by combining two seemingly disjoint technologies-biosynthetic display libraries and microfluidic systems. To isolate optimal ligands from these libraries, our project developed an automated magnetic bacterial cell sorting system, termed micromagnetic cell sorter (MMS), to screen microbial libraries with unprecedented throughput using a disposable microfluidic cartridge. We report on the sorting system performance in rare cell and ultra-rare cell recovery (populations less than 0.001%), and compare the results to the standard manual method, magnetic activated cell Sorting (MACS). To demonstrate the system effectiveness in reagent isolation through sorting against an anthrax toxin target, protective antigen was performed and comparable results to standard screening methods were obtained. In addition, the good recovery performance through MMS selection yielded a consensus sequence a...

Population Enrichment and Isolation with Magnetic Sorting

: Manual and semi-automated sorting methods (MACS) provide a high-throughput, less costly method ... more : Manual and semi-automated sorting methods (MACS) provide a high-throughput, less costly method for sample enrichment compared to fluorescence activated cell sorting (FACS). Magnetic sorting methods use commercially available magnetic beads with many surface modifications available for customizable sorting methods. In this note, we outline the methods for magnetic cell sorting using manual and benchtop magnets (MACS), as well as a micromagnetic sorter (MMS) that uses diposable, microfluidic cartridges. Along with magnetic sorting methods, we detail flow cytometry analysis techniques to quantify cell population enrichment for isolation of bacterial display library reagents.

Surface Assembly and Intracellular Delivery of Biomolecule Conjugated Nanomaterials

2.1 Introduction………………………………………………………………………….8 2.2 Methods and Results………………………………………………………….……9 ... more 2.1 Introduction………………………………………………………………………….8 2.2 Methods and Results………………………………………………………….……9 2.3 Discussion and Model….…….……………….………………………………...20 3. MAINTAINING STRUCTURE AND FUNCTION OF A HISTIDINE-TAGGED PROTEIN ATTACHED TO A 1.5 nm Au NANOPARTICLE…..……………………….23 3.1 Introduction………………………….……………………………………………..23 3.2 Materials and Methods……………………………………………………………25 3.3 Results…………………………………………………………………………..….28 3.4 Conclusions…………………………………………………………..……………38 4. DEVELOPMENT OF A BIMODAL MRI CONTRAST AGENT………………………...41 4.1 Introduction…………………………………..…………………………………….41 4.2 Materials and Methods….…………...………………………..……………….….46 4.3 Results……….………..…………………………………………………………....50 4.4 Conclusions……………………….…………………………………………..…...55 4.5 Future Work……………………………………………………………….……….56 5. NANOMEDIATED DELIVERY OF DUAL GENE REGULATORS…..………………..57 5.1 Introduction………………………………………………………………………...57 5.2 Materials and Methods……………………………………………………………59 v 5.3 Results………………………………………………………………………………61 5.4 Discussion………………………………………………………………………….71 REFERENCES…………………………………………………………………………………72 BIOGRAPHICAL SKETCH……………………………………………………….…..………87 vi LIST OF TABLES 2.1 FT-IR peak assignments of His 6-peptide and His 6 peptide with AuNP………….14 2.2 FT-IR peak assignments of Cys-peptide and Cys-peptide with AuNP………..…16 2.3 FT-IR peak assignments of Ser-peptide and Ser-peptide with AuNP……………17 3.1 FITC fluorescence recovery rates with FGF1 and FGF1-heparin………………..29 4.1 T1 and T2 relaxation rates for InP/ZnS-CAAKA-DOTA-Dy(III) cell layer experiments in agarose tissue mimic……………………………………………..…52 4.2 T1, T2, T2 linked, and T2 star results for InP/ZnS-CAAKA-DOTA-Dy(III) cell dosing experiments …………………………………………………………………..53 4.3 T1 and T2 relaxation rates for InP/ZnS-CAAKATat-DOTA-Dy(III) with and without cationic liposome transfecting agent………………………………………………..55 vii LIST OF FIGURES 1.1 Crystallographic model of 12 base pair duplex DNA and DsRed fluorescent protein…………………………………………………………………………………….3 1.2 Microscope image of CHO cells and TEM of 1.5 nm AuNP……….

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